ICN1-induced leukemia is extrathymic in origin. Bone marrow cells from BALB/c nu/nu mice were transduced with mICN1 or empty retroviruses and injected into BALB/c nu/nu recipients. Cell suspensions were stained for the expression of CD4 and CD8 markers, and FACS analysis was performed to determine EGFP⁺ and EGFP^- population percentages in peripheral blood (PB; E) 3 weeks after BMT and in bone marrow (BM; A) and spleen (SPL; C) 8 weeks after BMT of BALB/c nu/nu mice reconstituted with BALB/c nu/nu mice bone marrow transduced with mICN1-containing retroviruses. Percentage of CD4⁺CD8⁺ cells in the EGFP⁺ (B,D,F; R2, left panels) and EGFP^- (B,D,F; R3, right panels) fractions from the same samples are indicated. Total yield of spleen is also included. EGFP⁺CD4⁺CD8⁺ cells were never detected in peripheral blood and organs from BALB/c nu/nu mice that underwent reconstitution with BALB/c nu/nu mice bone marrow transduced with empty-containing retroviruses (data not shown). Data are representative of at least 3 independent experiments.